背景:过早卵巢功能不全(POI)是不孕症的主要原因。在这项研究中,我们旨在研究骨髓间充质干细胞(BMSCs)联合艾灸(BMSCs-MOX)对POI的影响,并评估其潜在机制。
方法:注射不同剂量的环磷酰胺(Cy)建立POI大鼠模型。通过评估发情周期来评估POI的建模和治疗的效果,血清激素水平,卵巢重量,卵巢指数,和卵巢组织病理学分析。通过追踪DiR标记的BMSCs,分析趋化因子基质细胞衍生因子1(Sdf1)和趋化因子受体4(Cxcr4)的表达,评价艾灸对BMSCs迁移的影响。通过测量活性氧(ROS)的水平来评估线粒体功能和线粒体自噬,线粒体膜电位(MMP),ATP,和线粒体自噬标记(Drp1,Pink1和Parkin)。此外,线粒体自噬抑制剂Mdivi-1和线粒体自噬激活剂CCCP用于证实线粒体自噬在Cy诱导的卵巢损伤中的作用以及联合治疗的潜在机制.
结果:使用Cy注射建立了合适的POI大鼠模型。与艾灸或BMSCs单独移植相比,BMSCs-MOX显示改善的结果,如减少发情周期障碍,改善卵巢重量和指数,正常的血清激素水平,卵巢储备增加,减少卵泡闭锁。艾灸增强了Sdf1和Cxcr4的表达,促进BMSCs迁移。BMSCs-MOX降低ROS水平;上调卵巢颗粒细胞(GC)中的MMP和ATP水平;并下调卵巢组织中Drp1,Pink1和Parkin的表达。Mdivi-1可显着减轻卵巢GCs中的线粒体功能障碍并改善卵巢功能。CCCP抑制BMSCs-MOX治疗调节线粒体自噬的能力,改善Cy诱导的卵巢损伤。
结论:艾灸可增强BMSCs移植后的迁移归巢能力,提高其修复卵巢损伤的能力。BMSCs联合艾灸可有效降低线粒体自噬的过度激活,有助于防止线粒体损伤,最终改善卵巢功能。这些发现为病理性卵巢衰老的治疗提供了新的方法,并为增强POI患者干细胞治疗的疗效提供了新的见解。
BACKGROUND: Premature ovarian insufficiency (POI) is a major cause of infertility. In this study, we aimed to investigate the effects of the combination of bone marrow mesenchymal stem cells (BMSCs) and moxibustion (BMSCs-MOX) on POI and evaluate the underlying mechanisms.
METHODS: A POI rat model was established by injecting different doses of cyclophosphamide (Cy). The modeling of POI and the effects of the treatments were assessed by evaluating estrous cycle, serum hormone levels, ovarian weight, ovarian index, and ovarian histopathological analysis. The effects of moxibustion on BMSCs migration were evaluated by tracking DiR-labeled BMSCs and analyzing the expression of chemokines stromal cell-derived factor 1 (Sdf1) and chemokine receptor type 4 (Cxcr4). Mitochondrial function and mitophagy were assessed by measuring the levels of reactive oxygen species (ROS), mitochondrial membrane potential (MMP), ATP, and the mitophagy markers (Drp1, Pink1, and Parkin). Furthermore, the mitophagy inhibitor Mdivi-1 and the mitophagy activator CCCP were used to confirm the role of mitophagy in Cy-induced ovarian injury and the underlying mechanism of combination therapy.
RESULTS: A suitable rat model of POI was established using Cy injection. Compared to moxibustion or BMSCs transplantation alone, BMSCs-MOX showed improved outcomes, such as reduced estrous cycle disorders, improved ovarian weight and index, normalized serum hormone levels, increased ovarian reserve, and reduced follicle atresia. Moxibustion enhanced Sdf1 and Cxcr4 expression, promoting BMSCs migration. BMSCs-MOX reduced ROS levels; upregulated MMP and ATP levels in ovarian granulosa cells (GCs); and downregulated Drp1, Pink1, and Parkin expression in ovarian tissues. Mdivi-1 significantly mitigated mitochondrial dysfunction in ovarian GCs and improved ovarian function. CCCP inhibited the ability of BMSCs-MOX treatment to regulate mitophagy and ameliorate Cy-induced ovarian injury.
CONCLUSIONS: Moxibustion enhanced the migration and homing of BMSCs following transplantation and improves their ability to repair ovarian damage. The combination of BMSCs and moxibustion effectively reduced the excessive activation of mitophagy, which helped prevent mitochondrial damage, ultimately improving ovarian function. These findings provide a novel approach for the treatment of pathological ovarian aging and offer new insights into enhancing the efficacy of stem cell therapy for POI patients.